A broad-spectrum chemokine antagonist encoded by Kaposi's sarcoma-associated herpesvirus.

Kaposi's sarcoma-associated herpesvirus encodes a chemokine called vMIP-II. This protein displayed a broader spectrum of receptor activities than any mammalian chemokine as it bound with high affinity to a number of both CC and CXC chemokine receptors. Binding of vMIP-II, however, was not associated with the normal, rapid mobilization of calcium from intracellular stores; instead, it blocked calcium mobilization induced by endogenous chemokines. In freshly isolated human monocytes the virally encoded vMIP-II acted as a potent and efficient antagonist of chemotaxis induced by chemokines. Because vMIP-II could inhibit cell entry of human immunodeficiency virus (HIV) mediated through CCR3 and CCR5 as well as CXCR4, this protein may serve as a lead for development of broad-spectrum anti-HIV agents.

Identification of Cys-150 in the active site of phosphomannose isomerase from Candida albicans.

Candida albicans phosphomannose isomerase (PMI) (EC 5.3.1.8) has been recently cloned and overexpressed in Escherichia coli. The enzyme can be irreversibly inactivated by iodoacetate in 50 mM borate buffer, pH 9.0, in a time-dependent manner at a rate of 4.2 +/- 0.03 min-1 M-1. This inhibition can be prevented by the substrate mannose 6-phosphate with a Ks of 0.22 +/- 0.05 mM, slightly lower than its Km value. However, metals such as zinc and cadmium, which are reversible, competitive inhibitors for PMI, do not protect the enzyme against modification. The protein has been labeled by using [2-14C]iodoacetate, in the presence or absence of substrate, and the protein is fully inactivated when 1.0 thiol group is modified per molecule of enzyme. Tryptic maps of the modified protein have been produced. The protected peptide has been identified and sequenced, and the phenylthiohydantoin amino acids have been collected. The modified amino acid is Cys-150. This cysteine residue is conserved in mammalian and yeast phosphomannose isomerases, but not in bacterial species where it is replaced with asparagine. We therefore purified PMI from E. coli and showed that this enzyme is not sensitive to inactivation by iodoacetate. The iodoacetate is presumably inhibiting PMI by sterically blocking the mannose 6-phosphate binding site. Multiple sequence alignment procedures were used to try to identify potential ligands of the zinc atom that is essential for enzyme activity and thus to delineate the active site region.(ABSTRACT TRUNCATED AT 250 WORDS)